Technical Field
Embodiments of the field of the disclosure include at least medical devices, medical diagnostics, immunology, cell biology, and molecular biology. In specific embodiments, the disclosure relates to the detection of analyte concentration in solutions prepared from bodily fluids or laboratory samples. The methods and devices in this disclosure are particularly useful for point-of-care medical and laboratory testing in a variety of medical or scientific applications.
Background of the Technology
The present invention relates to a method of detecting analyte concentration based on moving magnetic particles through multiple fluid environments, which are not in significant fluid communication, and are separated by gaseous separations. Specifically, the utility of this invention is to perform immunoassays or other diagnostic assays based on binding events in a rapid, simple, yet quantitative point-of-care testing device. A primary objective of the invention is to eliminate the time-consuming and labor intensive washing steps required to remove unreacted chemicals in conventional immunoassays by taking the reactants out of the solution using magnetic particles, effectively ‘washing off’ unreacted chemicals left behind in the solution.
In the field of healthcare, many testing devices and methods can be used to perform patient tests required by doctors to make diagnostic decisions. Determining the concentration of analytes in a solution is an essential quantitative parameter for many decisions made by doctors to improve patient outcome. In general, doctors inside or outside of a hospital environment work with other healthcare professionals in the laboratory environment to perform patient tests and determine key quantitative parameters by measuring the concentration of a variety of analytes in various bodily fluids. The immunoassay is one of the most commonly used laboratory tests for determining analyte concentration in the current medical and healthcare environment. Many healthcare situations require immediate determination of test results for the most efficacious treatment decisions. Unfortunately, immunoassays performed in a laboratory environment necessitate delays between sample collection and completion of the test. Point-of-care testing is preferable; as doctors or other healthcare professionals are able to produce rapid testing results for patient care decisions. Additionally, many healthcare environments lack dependable or available laboratory testing facilities or knowledge of laboratory testing methods, necessitating a point-of-care immunoassay device that is rapid, simple to use, and produces quantitative results. Beyond healthcare, immunoassays are required whenever it is necessary to measure the concentration of a protein or other macromolecule in solution. Examples include veterinary applications and detection of toxins in water for environmental monitoring.
Immunoassays are used to quantify the unknown concentration of an analyte within a sample. An immunoassay uses the selective binding of an antibody to the protein of interest to generate a signal and essentially comprises of (i) binding of the protein of interest to its corresponding antibody, and (ii) detecting the extent of this binding. The assays may be semi-quantitative or quantitative. There are two major varieties of immunoassays based on the number of antibodies binding to the analyte: sandwich and competitive. There are several methods to detect the extent of binding. These include enzymatic action on a substrate, fluorescent signals and radioactive signals. A conventional immunoassay, the enzyme linked immunosorbent assays (ELISA), is usually performed in microwell plates, such as 96-well plates).
Prior to detection, it is essential to remove unbound reactants. This is generally accomplished by serial washing (all sandwich ELISA variants). Washing increases the sensitivity of the detection. However, washing increases both the time and complexity of the operation, requiring either expert handling or significant automation cost and complexity. Another form of immunoassay is lateral flow assay (LFA) where the spontaneous flow of fluid along a chromatographic matrix is used to mix the different antibodies and analytes together. LFAs are typically small, quick, point-of-care devices. However, absence of washing during the flow limits the sensitivity and range of the detection process. Currently, there are no methods to do an accurate, quick, cheap, hand-held, point-of-use immunoassay.
Other assays can also be modified to be performed using the general principles of immunoassays. These include (but are not limited to) bioassays for cells (human cells, microorganisms, and other cell types), nucleic acids (mRNA, siRNA and others), and assays for other biomacromolecules.
The general principles of immunoassays may also be used for purposes of purification of biologics, including but not limited to the purification of proteins, DNA, RNA, lipids, or other biological molecules derived from complex mixtures and extracts of cells or tissues.
Magnetic particles are especially constructed to be attracted strongly to applied magnetic fields, but they retain no residual magnetism upon removal of the magnetic field. They are commonly used in immunoprecipitation—a method used to recover analytes of interest from a solution using repeated steps of magnetic precipitation, washing and elution.
Microfluidics is a discipline that deals with the behavior and manipulation of liquids that are geometrically constrained such that surface forces acting on the liquid, like surface tension and fluidic resistance dominate rather than bulk forces, like gravity, which predominate at the macro level. Microfluidics generally depend on the movement of liquid within the constrained spaces. Embodiments of the disclosure encompass methods and compositions that can be employed in a microfluidic form or in a larger fluidic embodiment.
US 2013/0206701 describes a method of transporting magnetic particles between separated fluid compartments in a rotational body that spins about a stationary magnet, using centrifugal forces and magnetic forces to alternate between fluid compartments and empty spaces within the rotational body. Disclosed therein is a rotating fluidic disc that contains both large empty spaces and fluidic regions and is utilized for the transport of magnetic particles through these regions. Presence in a region, though not control of location within a region, is enabled by rotation of the fluidic disc about a fixed magnet, with regions of the rotating fluidic disc designed to regulate particle movement. Currently, there exists no method for transporting magnetic particles between separated fluid compartments by movement of a magnet coupled to actuators relative to a fixed fluidic device as described in the present disclosure.
U.S. 2008/0073546 A1 describes methods of improved mixing by rotational movement in microfluidic structures in the context of magnetic particles located in fluid environments. Magnetic particle mixing patterns can be coarsely controlled by rotational frequency and rotational direction about a magnet.